Over 6,000 people require liver transplants every year in the Unites States alone. To date, several human liver transplant recipients have developed liver graft tolerance without immunosuppressive therapy, which suggests that the liver has tolerogenic properties. Kupffer Cells (KC), liver macrophages, have reported involvement in liver tolerance. Depletion of KC results in loss of tolerance and graft rejection in transplant models. Although KC play an important role in liver tolerance, the exact mechanisms behind liver tolerance remain unknown. Interestingly, gut bacteria are essential to the development of many aspects of the immune system. Additionally, bacteria of the intestinal flora influence the development of immune cells involved in liver tolerance. Because of the role of KC in liver tolerance, identifying the significance of gut bacteria in the development of these cells is an important goal. We hypothesize that the gut flora stimulates the development of the KC population and can modulate the tolerogenic potential of KC. Our preliminary data demonstrate a reduction in liver leukocytes in the absence of gut bacteria. Moreover, we observed a reduction in KC frequency in the absence of gut bacteria as well. This study will evaluate when gut bacteria influence KC development, the route of KC expansion promoted by gut bacteria, and the mechanism by which gut bacteria promote KC expansion. Moreover, we aim to determine the effect of gut bacteria on KC maturation, function, and tolerogenic potential. The results of this study will provide the foundation for future studies evaluating the therapeutic potential of gut flora manipulation to achieve liver tolerance via development of tolerogenic KC. PUBLIC HEALTH RELEVANCE: Liver transplantation outcome has drastically improved with immunosuppressive therapies like Cyclosporin A and Mycophenolate mofetil. However, many of the complications associated with liver transplantation (drug toxicity, infection, and malignancy) are caused by immunosuppressive therapy. By furthering our understanding of the mechanisms responsible for liver tolerance, identification of new strategies to achieve liver tolerance will be possible. Alternative strategies preventing liver graft rejection without suppressing the immune system have the potential to change the quality of life of thousands of people.